Thermoinduced structural changes in the membranes of animal organs following administration of antioxidants and during growth of malignancies

By means of weak-bound spin probes--stable iminoxyl free radicals differing in the level of hydrophobity there were studied thermoinduced structural transitions in the membranes of cell organells of animal liver after intraperitoneal injection of antioxidants and in the course of malignant growth, and in the nuclear membranes of Ehrlich ascite carcinoma as well. It has been found that during the growth of Ehrlich ascite carcinoma changes in rotationary mobility of probes in cell nuclei isolated from the liver of tumour-carrying animal are similar to the changes observed after antiocidants injection. A different pattern is observed in tumour cells. The membranes of ascite cell nuclei are characterized by a weak dependence of tau c on temperature for both probes. Within the temperature range studied no characteristic structural transitions proceeding in the nuclei of intact animals are observed.     

Lipid phase transitions measured in intact cells with Fourier transform infrared spectroscopy

Lipid phase transitions in membranes are thought to be a major damaging event during cooling of cells prior to cryopreservation or during warming after freeze-thaw has been completed. Although there is abundant evidence that such transitions occur in isolated phospholipids, the evidence that they are found in membranes in intact cells is less clear, due largely to technical difficulties in detecting such transitions in the complex mixtures of lipids and proteins found in natural membranes. We show here that Fourier transform infrared spectroscopy provides a rapid, convenient method for detecting these transitions in intact cells. We have used intact pollen grains of cattail (Typha latifolia) as a primary experimental subject. Spectra taken of the intact pollen grains show most of the features commonly seen in natural membrane vesicles or pure phospholipids. Shifts in the vibrational frequency and width of the CH2 bands with temperature can be used to detect lipid phase transitions. Biochemical analysis, coupled with the spectroscopy, was used to assign transitions to nonpolar and polar lipids. Finally, although assignment of the melting lipid unambiguously in other cells has not yet been made, we show that the transitions can nevertheless be detected in other intact cells, including those of four plant species and sperm of three animals.     

Epithelial to mesenchymal transition (EMT) of feto-maternal reproductive tissues generates inflammation: a detrimental factor for preterm birth

Human pregnancy is a delicate and complex process where multiorgan interactions between two independent systems, the mother, and her fetus, maintain pregnancy. Intercellular interactions that can define homeostasis at the various cellular level between the two systems allow uninterrupted fetal growth and development until delivery. Interactions are needed for tissue remodeling during pregnancy at both fetal and maternal tissue layers. One of the mechanisms that help tissue remodeling is via cellular transitions where epithelial cells undergo a cyclic transition from epithelial to mesenchymal (EMT) and back from mesenchymal to epithelial (MET). Two major pregnancy-associated tissue systems that use EMT, and MET are the fetal membrane (amniochorion) amnion epithelial layer and cervical epithelial cells and will be reviewed here. EMT is often associated with localized inflammation, and it is a well-balanced process to facilitate tissue remodeling. Cyclic transition processes are important because a terminal state or the static state of EMT can cause accumulation of proinflammatory mesenchymal cells in the matrix regions of these tissues and increase localized inflammation that can cause tissue damage. Interactions that determine homeostasis are often controlled by both endocrine and paracrine mediators. Pregnancy maintenance hormone progesterone and its receptors are critical for maintaining the balance between EMT and MET. Increased intrauterine oxidative stress at term can force a static (terminal) EMT and increase inflammation that are physiologic processes that destabilize homeostasis that maintain pregnancy to promote labor and delivery of the fetus. However, conditions that can produce an untimely increase in EMT and inflammation can be pathologic. These tissue damages are often associated with adverse pregnancy complications such as preterm prelabor rupture of the membranes (pPROM) and spontaneous preterm birth (PTB). Therefore, an understanding of the biomolecular processes that maintain cyclic EMT-MET is critical to reducing the risk of pPROM and PTB. Extracellular vesicles (exosomes of 40-160 nm) that can carry various cargo are involved in cellular transitions as paracrine mediators. Exosomes can carry a variety of biomolecules as cargo. Studies specifically using exosomes from cells undergone EMT can carry a pro-inflammatory cargo and in a paracrine fashion can modify the neighboring tissue environment to cause enhancement of uterine inflammation.     

INTERCELLULAR COMMUNICATION AND TISSUE GROWTH : II. Tissue Regeneration

Intercellular communication was examined in regenerating rat liver and urodele skin, two tissues of fast but normal growth. In both, cellular communication is in general as good as in their respective normal intact state. This stands in striking contrast to the lack of cellular communication in tissues with cancerous growth. Upon wounding of the urodele skin, the normally permeable junctional membranes of cells near the wound border seal themselves off, thereby insulating the interiors of the communicated cell systems from the exterior. When the cells of two opposing borders make mechanical contact in the course of wound closure, communication between them ensues within 30 min. Within this period all cell movement also ceases ("contact inhibition"). The possible implications of these findings in the control of tissue growth are discussed.     

Membrane model of the regulation of proliferation: the theory and interpretation of an experiment

The role of cell surface physical organization in the cell cycle regulation is analyzed within the framework of the earlier proposed theory (Chernavskii et al., 1982). Two models of cell surface are considered: hard-frame fluid-mosaic model (latticemosaic) and the fluid-mosaic one. The former deals with normal cells. The existence of integral carcasse or "frame" which is formed by the essential part of cross-linked membrane components and may have at least two different conformational states is hypothesized. The second model describes membranes of tumour cells. With the latter theory any mitogen (excluding the restoration of nutrient depletion) reduces the mechanical tensile strength of the frame and stimulates the general structural rearrangement of the plasma membrane. There are only two conformational transitions during the cell cycle which serve as signals for the beginning of S and M phases. If the values of tensile strength are great enough and therefore the conformational transitions are impossible, the cells pass into the resting (prereplicative--G01, or premitotical--G02) state. Three types of experiments are interpreted in the proposed theory: a) on differences in the action of growth factors on normal and tumour cell cycle, b) on the necessary condition for mitogenicity of lectins, c) on the stimulation of proliferation by mechanical deformation of cells.     

A common mechanism for the mechanosensitive regulation of apoptosis in different cell types and for different mechanical stimuli

In all kinds of tissue cells are influenced by mechanical forces. In vivo fibroblasts are exposed to mechanical tension and endothelial cells are subjected directly to hemodynamic flow. It has been shown that disturbance of the mechanical stimulus leads to apoptosis by induction of an autocrine loop with thrombospondin-1 as ligand and an integrin/integrin associated protein (CD47) complex as receptor. In the present study the nature of the mechanical stimulus has been exchanged for these two cell types. If fibroblasts are subjected to laminar flow apoptosis decreases about 20-fold whereas turbulence leads to an significant increase compared with the static conditions. If endothelial cells grown on thin silicone membranes are exposed to permanent and pulsatile uniaxial strain, the cells are completely devoid of apoptosis. The thrombospondin-1 secretion as well as the expression of CD47 occurs exclusively under mechanical relaxation respectively turbulence. So different types of cells seem to share a common sense deciding whether a mechanical stimulus induces or suppresses apoptosis and use a common molecular machinery for the regulation of the process.     

The effect of a phorbol ester on the lipid microviscosity of two endoplasmic reticulum membrane fractions isolated from Krebs II ascites cells.

This paper deals with microviscosity parameters and thermoinduced structural transitions in the lipids of smooth and heavy rough endoplasmic reticulum membranes isolated from Krebs II ascites cells incubated with the phorbol ester 12-O-tetradecanoyl-phorbol-13-acetate. The phorbol ester was found to bring about a threefold increase in the microviscosity of the lipids in heavy rough membranes. Spin probe I (2,2,6,6-tetrahydro-4-capryloyl-oxypiperidine-1-oxyl), localized in the surface layer of the membrane lipids, gave results which indicate an increased number of thermoinduced structural transitions in the smooth membranes in the treated cells due to the transitions occurring at relatively low temperature and a decreased number of such transitions in the heavy rough fraction especially at high temperature. For 5,6-benzo-2,2,4,4-tetramethyl-1,2,3,4-tetrahydro-gamma-carboline-oxyl, probe II, mainly distributed in the annular lipids, a decrease in the number of low temperature transitions in the smooth fraction was observed, while an increase occurred in the heavy rough one. The results obtained are discussed in terms of the effect of phorbol esters as promoters of tumor progression.     

Regulation of phosphoinositide breakdown by guanine nucleotides

Phosphoinositide hydrolysis is coupled to receptor systems involved in the elevation of cytosolic Ca2+ and activation of protein kinase C. In cell-free systems, guanine nucleotides are required to transduce the effects of receptor activation to phosphoinositide breakdown. Non-hydrolyzable guanine nucleotides stimulate phosphoinositide breakdown in permeabilized cells as well as membranes prepared from salivary glands, GH3 cells, neutrophils, hepatocytes and cerebral cortical tissue. In blowfly salivary gland membranes, 5-hydroxytryptamine stimulates a guanine-nucleotide dependent breakdown of both endogenous and exogenous phosphoinositide substrate through activation of phospholipase C. These data suggest that a GTP-binding protein modulates phospholipase C activity. The identity of this GTP-binding protein has not been established but may resemble other regulatory GTP-binding proteins which have been identified as transducing proteins in a variety of receptor systems.     

Comparing ion conductance recordings of synthetic lipid bilayers with cell membranes containing TRP channels

In this article we compare electrical conductance events from single channel recordings of three TRP channel proteins (TRPA1, TRPM2 and TRPM8) expressed in human embryonic kidney cells with channel events recorded on synthetic lipid membranes close to melting transitions. Ion channels from the TRP family are involved in a variety of sensory processes including thermo- and mechano-reception. Synthetic lipid membranes close to phase transitions display channel-like events that respond to stimuli related to changes in intensive thermodynamic variables such as pressure and temperature. TRP channel activity is characterized by typical patterns of current events dependent on the type of protein expressed. Synthetic lipid bilayers show a wide spectrum of electrical phenomena that are considered typical for the activity of protein ion channels. We find unitary currents, burst behavior, flickering, multistep-conductances, and spikes behavior in both preparations. Moreover, we report conductances and lifetimes for lipid channels as described for protein channels. Non-linear and asymmetric current–voltage relationships are seen in both systems. Without further knowledge of the recording conditions, no easy decision can be made whether short current traces originate from a channel protein or from a pure lipid membrane.     

Endothelial cell's biomechanical properties are regulated by invasive cancer cells

Most cancer-related deaths are caused by the ability of cancer cells to metastasize. This process includes the dissemination of cancer cells from the primary tumor side and their migration to targeted organ sites. During the migration of cancer cells through the connective tissue microenvironment, which consists of endothelial cells and extracellular matrix components, biomechanical properties are crucial for the efficiency and speed of cancer cell invasion and subsequently, metastases formation. Biomechanics can enable cancer cells to migrate through tissue, transmigrate through basement membranes as well as endothelial monolayers and form metastases in targeted organs. The current focus of cancer research still lies on the investigation of cancer cell's biochemical and molecular capabilities such as molecular genetics and gene signaling, but these approaches ignore the mechanical nature of the invasion process of cancer cells. Moreover, even the role of the endothelium during the transmigration and invasion of cells is not clear, it has been seen as a passive barrier, but this could not explain all novel findings. This review discusses how cancer cells alter the structural, biochemical and mechanical properties of the endothelium to regulate their own invasiveness through extracellular matrices and hence, through the tissue microenvironment. Finally, this review sheds light on the mechanical properties of cancer cells and the interacting endothelium and points out the importance of the mechanical properties as a critical determinant for the efficiency of cancer cell invasion and the overall progression of cancer. In conclusion, the regulation of the endothelial cell's biomechanical properties by cancer cells is a critical determinant of cancer cell invasiveness and may affect the future development of new cancer treatments.     

Current state and implications of research on biological effects of millimeter waves: A review of the literature

In recent years, research into biological and medical effects of millimeter waves (MMW) has expanded greatly. This paper analyzes general trends in the area and briefly reviews the most significant publications, proceeding from cell-free systems, dosimetry, and spectroscopy issues through cultured cells and isolated organs to animals and humans. The studies reviewed demonstrate effects of low-intensity MMW (10 mW/cm² and less) on cell growth and proliferation, activity of enzymes, state of cell genetic apparatus, function of excitable membranes, peripheral receptors, and other biological systems. In animals and humans, local MMW exposure stimulated tissue repair and regeneration, alleviated stress reactions, and facilitated recovery in a wide range of diseases (MMW therapy). Many reported MMW effects could not be readily explained by temperature changes during irradiation. The paper outlines some problems and uncertainties in the MMW research area, identifies tasks for future studies, and discusses possible implications for development of exposure safety criteria and guidelines. Bioelectromagnetics 19:393–413, 1998. © 1998 Wiley-Liss, Inc.     

Glycoconjugate pattern of membranes in the acinar cell of the rat pancreas

Lectin-binding studies were performed at the ultrastructural level to characterize glycoconjugate patterns on membrane systems in pancreatic acinar cells of the rat. Five lectins reacting with different sugar moieties were applied to ultrathin frozen sections: concanavalin A (ConA): glucose, mannose; wheat-germ agglutinin (WGA): N-acetylglucosamine, sialic acid; Ricinus communis agglutinin I (RCA I): galactose; Ulex europaeus agglutinin I (UEA I): L-fucose; soybean agglutinin (SBA): N-acetylgalactosamine). Binding sites of lectins were visualized either by direct conjugation to colloidal gold or by the use of a three-step procedure involving additional immune reactions. The rough endoplasmic reticulum and the nuclear envelope of acinar cells was selectively labelled for ConA. The membranes of the Golgi apparatus bound all lectins applied with an increasing intensity proceeding from the cis- to the trans-Golgi area for SBA, UEA I and WGA. In contrast RCA I selectively labelled the trans-Golgi cisternae. The membranes of condensing vacuoles and zymogen granules were labelled for all lectins used although the density of the label differed between the lectins. In contrast the content of zymogen granules failed to bind SBA and WGA. Lysosomal bodies (membranes and content) revealed binding sites for all lectins used. The plasma membranes were heavily labelled by all lectins except for SBA which showed only a weak binding to the lateral and the apical plasma membrane. These results are in accordance to current biochemical knowledge of the successive steps in the glycosylation of membrane proteins. It could be demonstrated, that the cryo-section technique is suitable for the fine structural localisation of surface glycoconjugates of plasma membranes and internal membranes in pancreatic acinar cells using plant lectins.     

Bioenergetics of life,disease and death phenomena

In this article, some new aspects of unified cell bioenergetics are presented. From the perspective of unified cell bioenergetics certain subsequent stages of cancer development, from initiation stage, through transformation to metastasis, are analyzed. Here we show that after transformation, cancer cells are permanently exposed to reactive oxygen species, that causes continual random DNA mutations and as a result genome and chromosomal destabilizations. The modern cancer attractor hypothesis has been extended in explaining cancer development. Discussion is conducted in light of current cancerogenesis research, including bioenergetic cancer initiation, the somatic mutation theory and the tissue organization field theory. In the article reasons complicating the discovery of patterns of cancer genome changes and cancer evolution are presented. In addition certain cancer therapeutic aspects are given attention to.     

A statistical mechanical model of the pre- and subtransitions of lecithin membranes

A statistical mechanical model with experimentally proved facts as starting points is presented. This model explains on molecular level, the pre- and subtransitions appearing in lipid membranes. The model describes the main features of the transitions, the hysteresis of the subtransition and the mobility changes of the heads and chains at these transitions. The model was expanded for phosphatidylcholine homologues with arbitrary chain lengths, and a qualitative agreement in the case of pretransition as far as a quantitative one for the subtransition were found.     

Polymeric membranes for guided bone regeneration

In this review, different barrier membranes for guided bone regeneration (GBR) are described as a useful surgical technique to enhance bone regeneration in damaged alveolar sites before performing implants and fitting other dental appliances. The GBR procedure encourages bone regeneration through cellular exclusion and avoids the invasion of epithelial and connective tissues that grow at the defective site instead of bone tissue. The barrier membrane should satisfy various properties, such as biocompatibility, non-immunogenicity, non-toxicity, and a degradation rate that is long enough to permit mechanical support during bone formation. Other characteristics such as tissue integration, nutrient transfer, space maintenance and manageability are also of interest. In this review, various non-resorbable and resorbable commercially available membranes are described, based on expanded polytetrafluoroethylene, poly(lactic acid), poly(glycolic acid) and their copolymers. The polyester-based membranes are biodegradable, permit a single-stage procedure, and have higher manageability than non-resorbable membranes; however, they have shown poor biocompatibility. In contrast, membranes based on natural materials, such as collagen, are biocompatible but are characterized by poor mechanical properties and stability due to their early degradation. Moreover, new approaches are described, such as the use of multi-layered, graft-copolymer-based and composite membranes containing osteoconductive ceramic fillers as alternatives to conventional membranes.     

The Biomechanical Properties of 3d Extracellular Matrices and Embedded Cells Regulate the Invasiveness of Cancer Cells

The malignancy of tumors depends on the biomechanical properties of cancer cells and their microenvironment, which enable cancer cells to migrate through the connective tissue, transmigrate through basement membranes and endothelial monolayers and form metastases in targeted organs. The current focus of cancer research is still based on biological capabilities such as molecular genetics and gene signaling, but these approaches ignore the mechanical nature of the invasion process of cancer cells. This review will focus on how structural, biochemical and mechanical properties of extracellular matrices (ECMs), and adjacent cells regulate the invasiveness of cancer cells. In addition, it presents how cancer cells create their own microenvironment by restructuring of the ECM and by interaction with stromal cells, which then further contribute to the progression of cancer disease. Finally, this review will point out that mechanical properties are a critical determinant for the efficiency of cancer cell invasion and the progression of cancer which might affect the future development of new cancer treatments.     

Candidates for the mechanosensory system in bone

Some potential mechanisms by which bone cells sense mechanical loads are described and hypotheses concerning the functioning of these mechanisms are explored. It is well known that bone tissue adapts its structure to its mechanical load environment. Recent research has illuminated the biological response of bone to mechanical loading at the cellular level, but the precise mechanosensory system that signals bone cells to deposit or resorb tissue has not been identified. The purpose of this paper is to describe the current status of this research and to suggest some possible mechanosensory systems by which bone cells might sense environmental loads.     

Micromachined interfaces: new approaches in cell immunoisolation and biomolecular separation

As a novel therapeutic application of microfabrication technology, a micromachined membrane-based biocapsule is described for the transplantation of protein-secreting cells without the need for immunosuppression. This new approach to cell encapsulation is based on microfabrication technology whereby immunoisolation membranes are bulk and surface micromachined to present uniform and well-controlled pore sizes as small as 10 nm, tailored surface chemistries, and precise microarchitecture. Through its ability to achieve highly controlled microarchitectures on size scales relevant to living systems (from microm to nm), microfabrication technology offers unique opportunities to more precisely engineer biocapsules that allow free exchange of the nutrients, waste products, and secreted therapeutic proteins between the host (patient) and implanted cells, but exclude lymphocytes and antibodies that may attack foreign cells. Microfabricated inorganic encapsulation devices may provide biocompatibility, in vivo chemical and mechanical stability, tailored pore geometries, and superior immunoisolation for encapsulated cells over conventional encapsulation approaches. By using microfabrication techniques, structures can be fabricated with spatial features from the sub-micron range up to several millimeters. These multi-scale structures correspond well with hierarchical biological structures, from proteins and sub-cellular organelles to the tissue and organ levels.     

Chemiosmotic systems in medicine

The concept of chemiosmotic systems arises from the pioneering work of Peter Mitchell on two fronts. One is concerned with the mechanisms by which molecules are transported across membranes which are generally barriers to such transport. These mechanisms are inevitably molecular, and are now yielding their secrets to a combination of structural protein chemistry and molecular biology. The other front is more physiological, and explores the functional relationships between metabolism and transport. Nevertheless, the two fronts form a continuum of mutally related structure and function. Chemiosmotic systems provide a hierarchy of complexity, starting from say a uniporter reconstituted in a chemically defined bilayer, and proceeding to greater complexity in mitochondria, chloroplasts, eukaryotic and prokaryotic cell membranes, and multicellular systems. Their relationship to medicine is profound, because they provide many opportunities for therapeutic intervention. In this paper I present an overview of chemiosmotic systems at different levels of complexity, both molecular and biological, of their involvements in pathology, and of possible pharmacological treatment or prevention of disease.     

Fibronectin in the microvasculature: localization in the pericyte-endothelial interstitium

The pericytes of capillaries are interesting cells which resemble the smooth muscle cells of larger vessels in some aspects of their morphology and behavior. In this report, their relationship to the underlying endothelium has been investigated in some detail. Using indirect, fluorescent immunocytochemical techniques on fresh and fixed tissues, it was found that fibronectin (an adhesive protein in many tissue culture systems) is concentrated in spots along vessels and is only faintly visible in the basement membranes of exhaustively perfused preparations. By electron microscopy, using a peroxidase immunocytochemical marker, these concentrations of fibronectin were seen to be localized to the pericyte-endothelial interstitia. Examination by TEM using a new fixation procedure demonstrated the organization of microfilaments and dense plaques along the pericyte membrane with fibrous and basement membrane-like material within this interstitial space. The arrangements of these elements suggest a mechanical linkage between the two cells. Such a linkage would allow contractions or relaxation of the pericyte to affect vessel diameter.